Identification of a novel EphB4 phosphodegron regulated by the autocrine IGFII/IRA axis in malignant mesothelioma

Our plain-English summary of Identification of a novel EphB4 phosphodegron regulated by the autocrine IGFII/IRA axis in malignant mesothelioma, by P. Scalia, G. Pandini, V. Carnevale, et al (Oncogene, 2019).

The study

• Authors: P. Scalia, G. Pandini, V. Carnevale, A. Giordano, S.J. Williams
• Journal: Oncogene
• Published: 2019;38(31):5987-6001
• Cited by: 15 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Malignant mesothelioma is a deadly disease with limited therapeutic options. EphB4 is an oncogenic tyrosine kinase receptor expressed in malignant mesothelioma as well as in a variety of cancers. It is involved in tumor microenvironment mediating angiogenesis and invasive cellular effects via both EphrinB2 ligand-dependent and independent mechanisms. The molecular network underlying EphB4 oncogenic effects is still unclear. Here we show that EphB4 expression in malignant mesothelioma cells is markedly decreased upon neutralization of cancer-secreted IGF-II. In particular, we demonstrate that EphB4 protein expression in malignant mesothelioma cells depend upon a degradation rescue mechanism controlled by the autocrine IGF-II-insulin receptor-A specific signaling axis. We show that the regulation of EphB4 expression is linked to a competing post-translational modification of its carboxy-terminal tail via phosphorylation of its tyrosine 987 by the Insulin receptor isoform-A kinase-associated activity in response to the autocrine IGF-II stimuli. Neutralization of this autocrine-induced EphB4-phosphorylation by IGF-II associates with the increased ubiquitination of EphB4 carboxy-terminal tail and with its rapid degradation. We also describe a novel Ubiquitin binding motif in the targeted region as part of the identified EphB4 phosphodegron and provide 3D modeling data supporting a possible model for the acute EphB4 PTM-driven regulation by IGF-II. Altogether, these findings disclose a novel molecular mechanism for the maintenance of EphB4-expression in malignant mesothelioma cells and other IGF-II-secreting cancers (IGF2omas).

In plain English

This was a laboratory study of cancer cells grown in culture, not of patients or asbestos exposure. The authors studied three malignant mesothelioma cell lines (NCI-H28, NCI-H2052 and MSTO-211H) that release a growth factor called IGF-II. Blocking that IGF-II with an antibody lowered the cells' level of EphB4, a protein linked to cancer growth, by between 54 and 82 percent. It also lowered a second protein, VEGF-A. In MSTO-211H cells the drop in EphB4 was fully reversible. Using engineered mouse cells, the authors found that IGF-II added a phosphate tag to EphB4 at tyrosine 987, acting through insulin receptor isoform A, not the IGF1 receptor. When IGF-II was blocked, EphB4 carried more ubiquitin, a tag the authors linked to the protein's breakdown: 2.7 to 4.8 times more in one test. A computer simulation suggested that the end of the protein becomes more exposed when tyrosine 987 is phosphorylated. The authors proposed this mechanism as a key checkpoint for gaining or maintaining a malignant phenotype in certain cancers. They stated that part of their model needs further experimental confirmation.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) study of cultured cancer cell lines and engineered mouse cells, with database and sequence searches and computer molecular dynamics modeling
• Who was studied: Three malignant mesothelioma cell lines (NCI-H28, NCI-H2052, MSTO-211H), plus mouse embryo fibroblasts lacking the igf1 receptor and engineered to express either human insulin receptor isoform A or human IGF1 receptor; other cancer cell lines were also profiled. No patients were studied; no place or dates are given.
• Exposure: No asbestos exposure was studied. The cells were treated with a neutralizing antibody against IGF-II, and the engineered mouse cells were stimulated with synthetic human IGF-II (10 nM and 50 nM). Cycloheximide was also used.
• Main results:
  • In the mesothelioma cell lines, the IGF-II-neutralizing antibody inhibited EphB4 protein expression by between 54 and 82 percent.
  • Blocking IGF-II also decreased VEGF-A levels in the same mesothelioma cell lines.
  • In MSTO211H cells, EphB4 levels recovered when the antibody block was removed; the authors described the effect as fully reversible.
  • In engineered mouse cells and a synthetic-peptide test, IGF-II increased tyrosine phosphorylation of EphB4 at tyrosine 987 through insulin receptor isoform A, but not through the IGF1 receptor.
  • In a test-tube ubiquitination assay using MSTO211H cell extracts, EphB4 ubiquitination increased 2.7 to 4.8 fold when the cells were deprived of IGF-II.
  • K63-type ubiquitination was the most prominent type found on EphB4 in MSTO211H cells.
  • A sequence search found that EphB4 region 958 to 974 shared 47% amino acid identity (76% positive score) with a motif in a ubiquitin-related protein (UBX-domain-containing protein 7).
  • In the computer simulation, phosphorylation of tyrosine 987 increased the exposed surface area of the protein's end region from about 1690 to 2250 square angstroms, more than 30%.
• The authors' conclusion: The authors proposed that this IGF-II-controlled mechanism, which keeps EphB4 from being broken down, is a key checkpoint for gaining or maintaining a malignant phenotype in certain cancers.
• Limitations the authors state:
  • The authors stated that their proposed model, in which IGF-II leads to phosphorylation at both tyrosine 987 and threonine 976, requires further experimental confirmation.
  • The authors stated that the possible interplay between hypoxia (low-oxygen culture conditions, with a medium pH of 6.9) and the IGF-II/EphB4 mechanism needs further study.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma.

Medical terms in this study, explained in our Lexicon: Malignant Mesothelioma, Tumor Microenvironment, Angiogenesis.

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• Data mining analysis of the PP2A cell cycle axis in mesothelioma patients — R. Pippa, S. Boffo, et al. (2020)
• Expression of the Wilms' tumor gene WT1 in human malignant mesothelioma cell lines and relationship to platelet-derived growth factor A and insulin-like growth factor 2 expression — A.W. Langerak, K.A. Williamson, et al. (1995)

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